Department of Biochemistry, Duke University School of Medicine, Durham, NC, USA.
Department of Chemistry, Duke University, Durham, NC, USA.
Nat Commun. 2021 Aug 31;12(1):5201. doi: 10.1038/s41467-021-25253-8.
N-methyladenosine (mA) is a post-transcriptional modification that controls gene expression by recruiting proteins to RNA sites. The modification also slows biochemical processes through mechanisms that are not understood. Using temperature-dependent (20°C-65°C) NMR relaxation dispersion, we show that mA pairs with uridine with the methylamino group in the anti conformation to form a Watson-Crick base pair that transiently exchanges on the millisecond timescale with a singly hydrogen-bonded low-populated (1%) mismatch-like conformation in which the methylamino group is syn. This ability to rapidly interchange between Watson-Crick or mismatch-like forms, combined with different syn:anti isomer preferences when paired (1:100) versus unpaired (10:1), explains how mA robustly slows duplex annealing without affecting melting at elevated temperatures via two pathways in which isomerization occurs before or after duplex annealing. Our model quantitatively predicts how mA reshapes the kinetic landscape of nucleic acid hybridization and conformational transitions, and provides an explanation for why the modification robustly slows diverse cellular processes.
N6-甲基腺苷（m6A）是一种转录后修饰，通过招募蛋白到 RNA 结合位点来控制基因表达。该修饰还通过尚未被理解的机制减缓生化过程。通过温度依赖（20°C-65°C）的 NMR 弛豫色散实验，我们发现 m6A 与尿嘧啶以反式构象结合，形成一个 Watson-Crick 碱基对，该碱基对在毫秒时间尺度上进行短暂交换，与一个单氢键结合的低占据（1%）类似错配的构象进行交换，其中甲基氨基呈顺式。这种在 Watson-Crick 或类似错配的构象之间快速交换的能力，以及配对时（1:100）与未配对时（10:1）不同的顺反异构偏好，解释了 m6A 如何在不影响高温下熔解的情况下，通过两种异构化发生在双链退火之前或之后的途径，稳健地减缓双链退火。我们的模型定量预测了 m6A 如何重塑核酸杂交和构象转变的动力学景观，并为为什么该修饰能够稳健地减缓各种细胞过程提供了解释。
